1. Field of the Invention
The present application relates to a process for preparation of leuprolide and its pharmaceutically acceptable salts.
2. Description of the Related Art
Leuprorelin or leuprolide acetate belongs to a family of drugs called gonadotropin-releasing hormone (GnRH) analogues. It is a synthetic nonapeptide analog which acts as an agonist at pituitary GnRH receptors. The chemical name is 5-oxo-L-prolyl-L-histidyl-L-tryptophyl-L-seryl-L-tyrosyl-D-leucyl-L-leucyl-L-arginyl-N-ethyl-L-prolinamide acetate (salt) and its primary sequence is Pyr-His-Trp-Ser-Tyr-D-Leu-Leu-Arg-Pro-NHEt. It may be utilized in the treatment of hormone-responsive cancer such as prostate or breast cancer, estrogen-dependent conditions, such as endometriosis or utrine fibroids, and to control precocious puberty. Leuprolide acetate is marketed as Viadur by Bayer AG, as Eligard by Sanofi-Aventis and as Lupron by Takeda Abbott Pharmaceuticals (TAP) in the United States.
Recently some syntheses utilized for preparing leuprolide or leuprolide acetate were disclosed in EP 2,119,725A1, WO 2008/44890A1 and WO 2011/148384A1.
European Patent Application EP 2,119,725A1 describes a method of preparing a peptide which is a C-terminal amide derivative, which comprises providing an amino acid attached in its C-terminal to a super-acid labile resin; coupling said amino acid with another amino acid in the presence of a coupling reagent; repeating the coupling step to obtain a peptide, wherein the peptide is protected with at least one protecting group which remains on the peptide upon its cleavage from the resin; cleaving said protected peptide from the resin by admixing with a mild acidic solution; and amidating the obtained protected peptide with a suitable amine.
FIG. 1 illustrates the route to leuprolide disclosed in EP 2,119,725A1. The preparation of leuprolide precursor [Pyr-His(Trt)-Trp-Ser(tBu)-Tyr(tBu)-D-Leu-Leu-Arg(Pbf)-Pro-OH] is cleaved from the resin by admixing a 1% TFA in DCM. Although the applicant emphasizes the process is performed under mild condition, the protecting groups of the leuprolide precursor and the super-acid labile resin cannot be removed to obtain the leuprolide precursor simultaneously. Thus, such a process inevitably needs to increase additional operations. Disadvantages of the process include resulting in a more tedious process of preparing leuprolide and other costly work-up procedures required at additional steps. Besides, since it is difficult to cleave the leuprolide precursor from a general acid-labile resin with a mild acidic solution, the process for preparing leuprolide is limited to the use of a super-acid labile resin. Furthermore, it is noted that the leuprolide precursor is prepared by coupling a crude peptide cleaved from the super acid-labile resin, and the coupling reaction involves the addition of TBTU/HOBT in the activation of carboxyl group of the peptide. However, HOBT is explosive and not suitable for performing the preparation. Therefore, the method does not have the excellent applicability in terms of the preparation of leuprolide.
Another route, disclosed in WO 20111148384A1 and shown in FIG. 2, which relates to a process for the preparation of leuprolide or its pharmaceutical acceptable salts thereof by synthesizing the peptide fragments by solid phase [7 and 5 amino acids fragment, i.e., Pyr-His(Trt)-Trp(Boc)-Ser(tBu)-D-Leu-Leu-OH and Pyr-His(Trt)-Trp(Boc)-Ser(tBu)-Tyr(tBu)-OH] and solution phase [2 and 4 amino acids fragment, i.e., H-Arg(Pbf)-Pro-NHEt and H-D-Leu-Leu-Arg(Pbf)-Pro-NHEt] respectively. The final solution phase condensation of these peptide fragments (7+2 and 5+4) led to a nonapeptide leuprolide in the protected form, i.e., Pyr-His(Trt)-Trp(Boc)-Ser(tBu)-D-Leu-Leu-Arg(Pbf)-Pro-NHEt. The protected nonapeptide leuprolide is deprotected to obtained leuprolide, and then leuprolide can be converted into pharmaceutically acceptable salts. Disadvantages of this processes include the synthesis of the materials H-Arg(Pbf)-Pro-NHEt and H-D-Leu-Leu-Arg(Pbf)-Pro-NHEt involves the use of the expensive material, i.e., Pro-NH—CH2CH3. Besides, the cleavage of the 7 and 5 amino acids fragment from the super acid-labile resin is carried out by using 1% TFA in DCM, which is similar to the method disclosed in EP 2,119,725A1. Therefore, there is the same deficiency that the protecting groups of the 7 and 5 amino acids fragment and the super-acid labile resin cannot be simultaneously removed to obtain a leuprolide precursor. Such a process inevitably needs to increase additional operations. Disadvantages of the process include resulting in a more tedious process of preparing leuprolide and other costly work-up procedures required at additional steps. As above, the process for preparing leuprolide is limited to the use of a super-acid labile resin but not suitable for using a general acid-labile resin. Furthermore, it is noted that the protected nonapeptide leuprolide is prepared by coupling the 7 and 5 amino acids fragment cleaved from the super acid-labile resin with the 2 and 4 amino acid fragments, and the coupling reaction involves the addition of HBTU/6-CI-HOBT/DIEA in the activation of carboxyl group of the peptide. However, actually HOBT is explosive and not suitable for performing the coupling reaction. Therefore, the method does not have the excellent applicability in terms of the preparation of leuprolide.
Yet another route, disclosed in WO 2008/44890A1 and shown in FIG. 3, which includes using a super acid-labile resin in the SPPS to prepare a leuprolide derivative, i.e., Pyr-His-Try-Ser(Bzl)-Tyr(Bzl)-D-Leu-Leu-Arg(NO2) which is obtained by adding weak acidic cleavage solution such as 2%-50% TFA/DCM, then the leuprolide precursor is reacted with Pro-NH—CH2CH3 to obtain Pyr-His-Try-Ser(Bzl)-Tyr(Bzl)-D-Leu-Leu-Arg(NO2)-Pro-NH—CH2CH3, and then the side-chain protecting groups are removed through catalytic hydrogen transfer reaction using by Pd/C and cyclohexadiene etc. Disadvantages of this process include the material Pro-NH—CH2CH3 is expensive and the hydrogenation reaction utilized for removing the side-chain protecting groups is also unsafe.
As such, there is a strong demand for a convenient process that is suitable for preparation of leuprolide or its pharmaceutically acceptable salts on a commercial scale with high purity and without complicated and costly purification steps. The present invention addresses this and other needs.